1. Field of the Invention
The invention relates to a method for the synthesis of materials in various formats containing surface-confined binding sites for solid phase synthesis products (e.g. peptides, oligonucleotides, oligosaccharides).
2. Background of the Invention
Molecular imprinting (G. Wulff, Angew. Chem., Int. Ed. Engl. 34 (1995) 1812-32) has emerged as a key technology in analytical and separation sciences (B. Sellergren (Ed.), Techniques and instrumentation in analytical chemistry, Vol. 23, Elsevier Science B.V., Amsterdam 2001; L. I.
Andersson, J. Chromatogr., B: Biomed. Sci. Appl. 745 (2000) 3-13; K. Haupt, K. Mosbach, Chem. Rev. 100 (2000) 2495-2504). The name refers to the synthesis of cross-linked polymers in the presence of templates, which may be small molecules, biological macromolecules, micro-organisms or crystals (B. Sellergren, Angew. Chem. Int. Ed. 39 (2000) 1031-1037).
The beauty of the molecular imprinting concept lies in its inherent simplicity. Functional monomers and the template to be imprinted form solution complexes which are subsequently incorporated into a cross-linked matrix upon polymerisation. Removal of the template leaves behind sites with a precise geometry and orientation of functional groups, allowing subsequent recognition of the template or a structurally-related compound. The molecularly imprinted polymer (MIP) thus created contains nanometer-sized binding sites in addition to larger sized pores (B. Sellergren, K. J. Shea, J. Chromatogr. 635 (1993) 31). Therefore, for guest molecules to access the host binding site they must penetrate pores, the size of which are difficult to control independently from the generation of the imprinted site. One way to decouple these processes is to immobilize the template on the surface of porous, disposable solids that act as molds to create a desired porosity (E. Yilmaz, K. Haupt, K. Mosbach, Angew. Chem., Int. Ed. 39 (2000) 2115-2118; M. M. Titirici, A. J. Hall, B. Sellergren, Chem. Mater. 14 (2002) 21-23).
In this way, the pore system is determined by the solid mold regardless of the conditions used to generate the imprinted sites. In addition, all imprinted sites are confined to the pore wall surface of the resulting material. Thus, access to these sites can be controlled by the porosity of the solid mold which may, in turn, allow substructures of larger target molecules to be recognised by the surface exposed sites. So far the feasibility of this approach has been demonstrated in the imprinting of small molecules, i.e. nucleotide bases (M. M. Titirici, A. J. Hall, B. Sellergren, Chem. Mater. 14 (2002) 21-23) and small drugs (E. Yilmaz, K. Haupt, K. Mosbach, Angew. Chem., Int. Ed. 39 (2000) 2115-2118).
Despite these advances, a thorough evaluation of the benefits of confining the sites to the pore wall surface is still lacking. Particularly lacking is any suggestion of how to use this concept for the development of affinity phases for the separation of biological macromolecules, e.g. peptides, proteins, oligo- or poly-nucleotides or oligo- or poly-saccharides (see for instance B. R. Hart, K. J. Shea, J. Am. Chem. Soc. 123 (2001) 2072-2073; A. Rachkov, N. Minoura, Biochim. Biophys. Acta 1544 (2001) 255-266 for other examples of imprinted peptide receptors). In this regard, the format would allow a more efficient exploitation of the epitope approach, recently introduced by Rachkov and Minoura (A. Rachkov, N. Minoura, Biochim. Biophys. Acta 1544 (2001) 255-266). In this approach, a smaller peptide corresponding to a unique amino acid sequence of a target protein is used as template in order to generate a site that can subsequently selectively bind the larger target molecule. This requires that the site is associated with the accessible surface of larger pores capable of accommodating the larger protein.